The active protein component Yt from macrofungi Agrocybe aegerita has been found to have antitumor and immune regulation activity. For the investigation of the proteins, a cDNA library of the Agrocybe aegerita fruiting body was constructed and screened. The cDNA of a novel fungal Ub-conjugating enzyme (UBC) was identified (Genbank accession code: JQ362395), and sequence analysis revealed that it contained the conserved residues and belonged to the UBC family. Recombinant UBC was expressed using the pET system and purified, and obtained 10 mg from 1 L E coli. culture media. Exogenously applied UBC (1.8, 3.6 and 7.2 μM) in cell cultures altered HeLa cell morphology and induced cell apoptosis determined by flow cytometry, and the percentage of AnnexinV+/PI- cells was increased from 1.4% in control to 10.7%, 15.8% and 20.3%, which was concentration dependent. This is the first report on the effects of a fungal UBC on cancer cells, which suggests that the members of the fungal ubiquitin system might alter fundamental cellular processes and require further investigation.